Covalent diabodies and uses thereof

Abstract

The present invention is directed to diabody molecules and uses thereof in the treatment of a variety of diseases and disorders, including immunological disorders, infectious disease, intoxication and cancers. The diabody molecules of the invention comprise two polypeptide chains that associate to form at least two epitope binding sites, which may recognize the same or different epitopes on the same or differing antigens. Additionally, the antigens may be from the same or different molecules. The individual polypeptide chains of the diabody molecule may be covalently bound through nonpeptide bond covalent bonds, such as, but not limited to, disulfide bonding of cysteine residues located within each polypeptide chain. In particular embodiments, the diabody molecules of the present invention further comprise an Fc region, which allows antibody like functionality to engineered into the molecule.

What is claimed: 
     
       1. A diabody molecule capable of binding to an epitope of CD32B and an epitope of CD79B, wherein said diabody comprises a first polypeptide chain and a second polypeptide chain, said polypeptide chains each having an N-terminal end and a C-terminal end and being covalently bonded to one another, wherein:
 (A) said first polypeptide chain comprises, in the N-terminal to C-terminal direction:
 (i) a first domain comprising a binding region of a light chain variable domain of a first immunoglobulin (VL1) specific for binding said epitope of CD79B; 
 (ii) a second domain comprising a binding region of a heavy chain variable domain of a second immunoglobulin (VH2) specific for binding said epitope of CD32B; and 
 (iii) a third domain comprising at least one cysteine residue; 
 wherein said first domain and said second domain are linked to one another by an intervening linker such that said first domain and said second domain are constrained from self-assembly and do not associate to form an epitope binding site; and 
 
 (B) said second polypeptide chain comprises, in the N-terminal to C-terminal direction:
 (i) a fourth domain comprising a binding region of a light chain variable domain of said second immunoglobulin (VL2) specific for binding said epitope of CD32B; 
 (ii) a fifth domain comprising a binding region of a heavy chain variable domain of said first immunoglobulin (VH1) specific for binding CD79B; and 
 (iii) a sixth domain comprising at least one cysteine residue; 
 wherein said fourth domain and said fifth domain are linked to one another by an intervening linker such that said fourth domain and said fifth domain are constrained from self-assembly and do not associate to form an epitope binding site; 
 
 wherein: 
 (1) said first domain and said fifth domain are associated to form a binding site (VL1/VH1) that binds said epitope of CD79B, said second domain and said fourth domain associate to form a binding site (VH2/VL2) that binds said epitope of CD32B, and said cysteine residues of said third domain and said sixth domain form a disulfide bond between said first and second polypeptide chains; 
 (2) said first polypeptide chain comprises the light chain variable domain, the intervening GGGSGGGG linker (SEQ ID NO:10) and the heavy chain variable domain of SEQ ID NO:132; and 
 (3) said second polypeptide chain comprises:
 (i) the light chain variable domain, the intervening GGGSGGGG linker (SEQ ID NO:10) and the heavy chain variable domain of SEQ ID NO:134; or 
 (ii) the light chain variable domain, the intervening GGGSGGGG linker (SEQ ID NO:10) and the heavy chain variable domain of SEQ ID NO:136. 
 
 
     
     
       2. The diabody molecule of  claim 1 , wherein said second polypeptide chain comprises said light chain variable domain, said intervening GGGSGGGG linker (SEQ ID NO:10) and said heavy chain variable domain of SEQ ID NO:134. 
     
     
       3. The diabody molecule of  claim 1 , wherein said second polypeptide chain comprises said light chain variable domain, said intervening GGGSGGGG linker (SEQ ID NO:10) and said heavy chain variable domain of SEQ ID NO:136. 
     
     
       4. A pharmaceutical composition comprising a therapeutically effective amount of the diabody molecule of  claim 1 , and a pharmaceutically acceptable carrier. 
     
     
       5. The diabody molecule of  claim 1 , wherein said first polypeptide chain or said second polypeptide chain further comprises an Fc domain or portion thereof. 
     
     
       6. The diabody molecule of  claim 5 , wherein said Fc domain or portion thereof is linked to the N-terminus of said first domain or said fourth domain. 
     
     
       7. The diabody molecule of  claim 1 , wherein said first polypeptide chain or said second polypeptide chain further comprises an Fc domain or portion thereof. 
     
     
       8. The diabody molecule of  claim 7 , wherein said Fc domain or portion thereof is linked to the N-terminus of said first domain or said fourth domain. 
     
     
       9. A pharmaceutical composition comprising a therapeutically effective amount of the diabody molecule of  claim 1 , and a pharmaceutically acceptable carrier.